Publications by authors named "Mitchell Levesque"

136 Publications

Cooperation between melanoma cell states promotes metastasis through heterotypic cluster formation.

Dev Cell 2021 Sep 13. Epub 2021 Sep 13.

Cancer Biology and Genetics, Memorial Sloan Kettering Cancer Center, New York, NY 10065, USA. Electronic address:

Melanomas can have multiple coexisting cell states, including proliferative (PRO) versus invasive (INV) subpopulations that represent a "go or grow" trade-off; however, how these populations interact is poorly understood. Using a combination of zebrafish modeling and analysis of patient samples, we show that INV and PRO cells form spatially structured heterotypic clusters and cooperate in the seeding of metastasis, maintaining cell state heterogeneity. INV cells adhere tightly to each other and form clusters with a rim of PRO cells. Intravital imaging demonstrated cooperation in which INV cells facilitate dissemination of less metastatic PRO cells. We identified the TFAP2 neural crest transcription factor as a master regulator of clustering and PRO/INV states. Isolation of clusters from patients with metastatic melanoma revealed a subset with heterotypic PRO-INV clusters. Our data suggest a framework for the co-existence of these two divergent cell populations, in which heterotypic clusters promote metastasis via cell-cell cooperation.
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http://dx.doi.org/10.1016/j.devcel.2021.08.018DOI Listing
September 2021

Real-life data for first-line combination immune-checkpoint inhibition and targeted therapy in patients with melanoma brain metastases.

Eur J Cancer 2021 Aug 25;156:149-163. Epub 2021 Aug 25.

Department of Dermatology, University Hospital Zurich, Zurich, Switzerland.

Background: Melanoma brain metastases (MBM) have a poor prognosis. Systemic treatments that have improved outcomes in advanced melanoma have been shown to have an intracranial (IC) effect. We studied the efficacy and outcomes of combined immune checkpoint inhibitor ipilimumab/nivolumab (Combi-ICI) or targeted therapy (Combi-TT) as first-line treatment in MBM.

Methods: MBM patients treated with Combi-ICI or Combi-TT within 3 months after MBM diagnosis. Endpoints were progression-free survival (PFS) and overall survival (OS).

Results: 53 patients received Combi-ICI, 32% had symptomatic MBM and 33.9% elevated LDH. 71.7% required local treatment. The disease control rate was 60.3%. IC response rate (RR) was 43.8% at 3-months with durable responses at 6- (46.5%) and 12-months (53.1%). Extracranial (EC) RR was 44.7% at 3-months and 50% at 12-months. Median PFS was 9.6 months (95% CI 3.6-NR) and median overall survival (mOS) 44.8 months (95% CI; 26.2-NR). 63 patients received Combi-TT, 55.6% of patients had symptomatic MBM, 57.2% of patients had elevated LDH and 68.3% of patients required local treatment. The disease control rate was 60.4%. ICRR was 50% at 3-months, but dropped at 6-months (20.9%). ECRR was 69.2% at 3-months and 17.6% at 12-months. Median PFS was 5.8 months (95% CI 4.2-7.6) and mOS 14.2 months (95% CI 8.99-26.8). In BRAFV600 patients, 26.7% of patients received Combi-ICI and 73.3% Combi-TT with OS (p = 0.0053) and mPFS (p = 0.03) in favour to Combi-ICI.

Conclusion: Combi-ICI showed prolonged mOS with sustainable IC and EC responses. Despite the initially increased efficacy, Combi-TT responses at 12 months were low. Combi-ICI appeared superior to Combi-TT for OS and PFS in BRAFV600 patients. Other clinical factors are determinants for first-line treatment choice.
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http://dx.doi.org/10.1016/j.ejca.2021.07.028DOI Listing
August 2021

Establishing standardized immune phenotyping of metastatic melanoma by digital pathology.

Lab Invest 2021 Aug 26. Epub 2021 Aug 26.

Department of Pathology and Molecular Pathology, University and University Hospital Zurich, Zurich, Switzerland.

CD8+ tumor-infiltrating T cells can be regarded as one of the most relevant predictive biomarkers in immune-oncology. Highly infiltrated tumors, referred to as inflamed (clinically "hot"), show the most favorable response to immune checkpoint inhibitors in contrast to tumors with a scarce immune infiltrate called immune desert or excluded (clinically "cold"). Nevertheless, quantitative and reproducible methods examining their prevalence within tumors are lacking. We therefore established a computational diagnostic algorithm to quantitatively measure spatial densities of tumor-infiltrating CD8+ T cells by digital pathology within the three known tumor compartments as recommended by the International Immuno-Oncology Biomarker Working Group in 116 prospective metastatic melanomas of the Swiss Tumor Profiler cohort. Workflow robustness was confirmed in 33 samples of an independent retrospective validation cohort. The introduction of the intratumoral tumor center compartment proved to be most relevant for establishing an immune diagnosis in metastatic disease, independent of metastatic site. Cut-off values for reproducible classification were defined and successfully assigned densities into the respective immune diagnostic category in the validation cohort with high sensitivity, specificity, and precision. We provide a robust diagnostic algorithm based on intratumoral and stromal CD8+ T-cell densities in the tumor center compartment that translates spatial densities of tumor-infiltrating CD8+ T cells into the clinically relevant immune diagnostic categories "inflamed", "excluded", and "desert". The consideration of the intratumoral tumor center compartment allows immune phenotyping in the clinically highly relevant setting of metastatic lesions, even if the invasive margin compartment is not captured in biopsy material.
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http://dx.doi.org/10.1038/s41374-021-00653-yDOI Listing
August 2021

Microgreens: Functional Food with Antiproliferative Cancer Properties Influenced by Light.

Foods 2021 Jul 22;10(8). Epub 2021 Jul 22.

Department of Agricultural and Food Sciences, University of Bologna, Viale Fanin, 44-40127 Bologna, Italy.

The anti-proliferative/pro-oxidant efficacy of green pea, soybean, radish, Red Rambo radish, and rocket microgreens, cultivated under either fluorescent lighting (predominant spectral peaks in green and orange) or combination light-emitting diode (LED, predominant spectral peak in blue) was investigated using Ewing sarcoma lines, RD-ES and A673, respectively. All aqueous microgreen extracts significantly reduced cell proliferation (cancer prevention effect) to varying extents in two-dimensional sarcoma cell cultures. The effect of the polyphenol fraction in the aqueous food matrix was unrelated to total polyphenol content, which differed between species and light treatment. Only (LED-grown) extracts exercised anti-proliferative and pro-apoptotic effects in both three-dimensional RD-ES and A673 spheroids (early tumor progression prevention), without cytotoxic effects on healthy L929 fibroblasts. A similar anti-tumor effect of Red Rambo radish (LED and fluorescent-grown) was evident only in the RD-ES spheroids. Aside from the promising anti-tumor potential of the polyphenol fraction of green pea microgreens, the latter also displayed favorable growth quality parameters, along with radish, under both light treatments over the 10 day cultivation period.
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http://dx.doi.org/10.3390/foods10081690DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8392261PMC
July 2021

Epigenetic control of melanoma cell invasiveness by the stem cell factor SALL4.

Nat Commun 2021 08 20;12(1):5056. Epub 2021 Aug 20.

University of Zürich, Institute of Anatomy, Zürich, Switzerland.

Melanoma cells rely on developmental programs during tumor initiation and progression. Here we show that the embryonic stem cell (ESC) factor Sall4 is re-expressed in the Tyr::Nras; Cdkn2a melanoma model and that its expression is necessary for primary melanoma formation. Surprisingly, while Sall4 loss prevents tumor formation, it promotes micrometastases to distant organs in this melanoma-prone mouse model. Transcriptional profiling and in vitro assays using human melanoma cells demonstrate that SALL4 loss induces a phenotype switch and the acquisition of an invasive phenotype. We show that SALL4 negatively regulates invasiveness through interaction with the histone deacetylase (HDAC) 2 and direct co-binding to a set of invasiveness genes. Consequently, SALL4 knock down, as well as HDAC inhibition, promote the expression of an invasive signature, while inhibition of histone acetylation partially reverts the invasiveness program induced by SALL4 loss. Thus, SALL4 appears to regulate phenotype switching in melanoma through an HDAC2-mediated mechanism.
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http://dx.doi.org/10.1038/s41467-021-25326-8DOI Listing
August 2021

Frequency, Treatment and Outcome of Immune-Related Toxicities in Patients with Immune-Checkpoint Inhibitors for Advanced Melanoma: Results from an Institutional Database Analysis.

Cancers (Basel) 2021 Jun 11;13(12). Epub 2021 Jun 11.

Department of Dermatology, University Hospital of Zurich, 8091 Zurich, Switzerland.

Immune checkpoint inhibitors (ICIs) can induce immune-related adverse events (irAEs), which may result in treatment discontinuation. We sought to describe the onset, frequency, and kinetics of irAEs in melanoma patients in a real-life setting and to further investigate the prognostic role of irAEs in treatment outcomes. In this retrospective single-center cohort study, we included 249 melanoma patients. Onset, grade, and resolution of irAEs and their treatment were analyzed. A total of 191 (74.6%) patients in the non-adjuvant and 65 (25.3%) in the adjuvant treatment setting were identified. In the non-adjuvant setting, 29 patients (59.2%) with anti-CTLA4, 43 (58.1%) with anti-PD1, and 54 (79.4%) with anti-PD1/anti-CTLA4 experienced some grade of irAE and these had an improved outcome. In the adjuvant setting, the frequency of irAEs was 84.6% in anti-CTLA4 and 63.5% in anti-PD1, but no correlation with disease relapse was observed. Patients with underlying autoimmune conditions have a risk of disease exacerbation. Immunomodulatory agents had no impact on treatment efficacy. IrAEs are correlated with increased treatment efficacy in the non-adjuvant setting. Application of steroids and immunomodulatory agents, such as anti-TNF-alpha or anti-IL6, did not affect ICI efficacy. These data support irAEs as possible prognostic markers for ICI treatment.
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http://dx.doi.org/10.3390/cancers13122931DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8230729PMC
June 2021

Clinicopathological and Genomic Profiles of Atypical Fibroxanthoma and Pleomorphic Dermal Sarcoma Identify Overlapping Signatures with a High Mutational Burden.

Genes (Basel) 2021 Jun 25;12(7). Epub 2021 Jun 25.

Dermatology Department, University Hospital Zurich, 8091 Zurich, Switzerland.

Atypical fibroxanthoma (AFX) and pleomorphic dermal sarcoma (PDS) are rare tumors developing in chronically sun-exposed skin. Clinicopathological features are similar, but they differ in prognosis, while PDS has a more aggressive course with a higher risk for local recurrence and metastases. In current clinical practice, they are diagnosed by exclusion using immunohistochemistry. Thus, stringent diagnostic criteria and correct differentiation are critical in management and treatment for optimal outcomes. This retrospective single-center study collected clinicopathological data and tumor samples of 10 AFX and 18 PDS. Extracted genomic DNA from tumor specimens was analyzed by a next-generation sequencing (NGS) platform (FoundationOne-CDx™). Among 65 identified mutations, inactivating mutations were observed in all tumor specimens. In both AFX and PDS, the known pathogenic gene alterations in , promoter, and were frequently present, along with high mutational burden and stable Micro-Satellite Instability status. The mutational profiles differed only in , which was only present in AFX. Further differences were identified in likely pathogenic and unknown gene alterations. Similarities in their genomic signatures could help to distinguish them from other malignancies, but they are not distinguishable between each other using the FoundationOne-CDx™ NGS panel. Therefore, histological criteria to determine diagnosis remain valid. For further insight, performing deep tumor profiling may be necessary.
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http://dx.doi.org/10.3390/genes12070974DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8303615PMC
June 2021

Cutaneous and systemic hyperinflammation drives maculopapular drug exanthema in severely ill COVID-19 patients.

Allergy 2021 Jun 22. Epub 2021 Jun 22.

Department of Dermatology, University Hospital Zurich, Zurich, Switzerland.

Background: Coronavirus disease-2019 (COVID-19) has been associated with cutaneous findings, some being the result of drug hypersensitivity reactions such as maculopapular drug rashes (MDR). The aim of this study was to investigate whether COVID-19 may impact the development of the MDR.

Methods: Blood and skin samples from COVID-19 patients (based on a positive nasopharyngeal PCR) suffering from MDR (COVID-MDR), healthy controls, non-COVID-19-related patients with drug rash with eosinophilia and systemic symptoms (DRESS), and MDR were analyzed. We utilized imaging mass cytometry (IMC) to characterize the cellular infiltrate in skin biopsies. Furthermore, RNA sequencing transcriptome of skin biopsy samples and high-throughput multiplexed proteomic profiling of serum were performed.

Results: IMC revealed by clustering analyses a more prominent, phenotypically shifted cytotoxic CD8 T cell population and highly activated monocyte/macrophage (Mo/Mac) clusters in COVID-MDR. The RNA sequencing transcriptome demonstrated a more robust cytotoxic response in COVID-MDR skin. However, severe acute respiratory syndrome coronavirus 2 was not detected in skin biopsies at the time point of MDR diagnosis. Serum proteomic profiling of COVID-MDR patients revealed upregulation of various inflammatory mediators (IL-4, IL-5, IL-6, TNF, and IFN-γ), eosinophil and Mo/Mac -attracting chemokines (MCP-2, MCP-3, MCP-4 and CCL11). Proteomics analyses demonstrated a massive systemic cytokine storm in COVID-MDR compared with the relatively milder cytokine storm observed in DRESS, while MDR did not exhibit such features.

Conclusion: A systemic cytokine storm may promote activation of Mo/Mac and cytotoxic CD8 T cells in severe COVID-19 patients, which in turn may impact the development of MDR.
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http://dx.doi.org/10.1111/all.14983DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8441838PMC
June 2021

Collection and preprocessing of fine needle aspirate patient samples for single cell profiling and data analysis.

STAR Protoc 2021 Jun 5;2(2):100581. Epub 2021 Jun 5.

Dermatology Department, University Hospital Zurich and Medical Faculty, University of Zurich, 8091 Zurich, Switzerland.

High cell viability and recovered cell concentration are typical quality control requirements for single-cell processing and quality data. This protocol describes procedures for sampling, live-cell biobanking, preprocessing for single-cell RNA sequencing, and analysis of fine-needle aspiration (FNA) samples of the skin. The minimally invasive nature of FNA collection is more accepted by patients and allows for frequent longitudinal sampling, resulting in high-quality single-cell sequencing data that capture cellular heterogeneity in clinical samples.
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http://dx.doi.org/10.1016/j.xpro.2021.100581DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8190505PMC
June 2021

Evolutionary predictability of genetic versus nongenetic resistance to anticancer drugs in melanoma.

Cancer Cell 2021 Aug 17;39(8):1135-1149.e8. Epub 2021 Jun 17.

Laboratory for Molecular Cancer Biology, Center for Cancer Biology, VIB, Leuven, Belgium; Laboratory for Molecular Cancer Biology, Department of Oncology, KU Leuven, Leuven, Belgium. Electronic address:

Therapy resistance arises from heterogeneous drug-tolerant persister cells or minimal residual disease (MRD) through genetic and nongenetic mechanisms. A key question is whether specific molecular features of the MRD ecosystem determine which of these two distinct trajectories will eventually prevail. We show that, in melanoma exposed to mitogen-activated protein kinase therapeutics, emergence of a transient neural crest stem cell (NCSC) population in MRD concurs with the development of nongenetic resistance. This increase relies on a glial cell line-derived neurotrophic factor-dependent signaling cascade, which activates the AKT survival pathway in a focal adhesion kinase (FAK)-dependent manner. Ablation of the NCSC population through FAK inhibition delays relapse in patient-derived tumor xenografts. Strikingly, all tumors that ultimately escape this treatment exhibit resistance-conferring genetic alterations and increased sensitivity to extracellular signal-regulated kinase inhibition. These findings identify an approach that abrogates the nongenetic resistance trajectory in melanoma and demonstrate that the cellular composition of MRD deterministically imposes distinct drug resistance evolutionary paths.
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http://dx.doi.org/10.1016/j.ccell.2021.05.015DOI Listing
August 2021

Who's Driving? Switch of Drivers in Immunotherapy-Treated Progressing Sinonasal Melanoma.

Cancers (Basel) 2021 May 31;13(11). Epub 2021 May 31.

Department of Pathology and Molecular Pathology, University Hospital Zurich, 8091 Zurich, Switzerland.

Mucosal melanoma can be driven by various driver mutations in genes such as , , or . However, some cases present with only weak drivers, or lacking known oncogenic drivers, suggesting immunotherapy over targeted therapy. While resistance mechanisms to immunotherapy in cutaneous melanoma have been uncovered, including alterations in /, or , a switch of oncogenic drivers under immunotherapy has not yet been observed. We report three cases of metastatic sinonasal melanoma that switched oncogenic drivers from , or no driver to during or after immunotherapy, thereby showing progressive disease. One of the cases presented with three spatially separate driver mutations in the primary tumor, whereas the clone persisted under immunotherapy. In comparison, three different control cases receiving radiotherapy only did not show a change of the detectable molecular drivers in their respective recurrences or metastases. In summary, these data provide an important rationale for longitudinal molecular testing, based on evidence for an unforeseen recurrent event of molecular driver switch to in progressing sinonasal melanoma. These findings provide the basis for further studies on a potential causal relation of emerging mutant clones and immunotherapy.
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http://dx.doi.org/10.3390/cancers13112725DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8198298PMC
May 2021

A Comparative Study of Real-Time RT-PCR-Based SARS-CoV-2 Detection Methods and Its Application to Human-Derived and Surface Swabbed Material.

J Mol Diagn 2021 07 5;23(7):796-804. Epub 2021 May 5.

Department of Dermatology, University Hospital Zurich, University of Zurich, Zurich, Switzerland. Electronic address:

Real-time RT-PCR remains a gold standard in the detection of various viral diseases. In the coronavirus 2019 pandemic, multiple RT-PCR-based tests were developed to screen for viral infection. As an emergency response to increasing testing demand, we established a severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) PCR diagnostics platform for which we compared different commercial and in-house RT-PCR protocols. Four commercial, one customized, and one in-house RT-PCR protocols were evaluated with 92 SARS-CoV-2-positive and 92 SARS-CoV-2-negative samples. Furthermore, economical and practical characteristics of these protocols were compared. In addition, a highly sensitive digital droplet PCR (ddPCR) method was developed, and application of RT-PCR and ddPCR methods on SARS-CoV-2 environmental samples was examined. Very low limits of detection (1 or 2 viral copies/μL), high sensitivities (93.6% to 97.8%), and high specificities (98.7% to 100%) for the tested RT-PCR protocols were found. Furthermore, the feasibility of downscaling two of the commercial protocols, which could optimize testing capacity, was demonstrated. Tested commercial and customized RT-PCR detection kits show very good and comparable sensitivity and specificity, and the kits could be further optimized for use on SARS-CoV-2 viral samples derived from human and surface swabbed samples.
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http://dx.doi.org/10.1016/j.jmoldx.2021.04.009DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8096526PMC
July 2021

Prolonged Unfrozen Storage and Repeated Freeze-Thawing of SARS-CoV-2 Patient Samples Have Minor Effects on SARS-CoV-2 Detectability by RT-PCR.

J Mol Diagn 2021 06 26;23(6):691-697. Epub 2021 Mar 26.

Department of Dermatology, University Hospital Zurich and Medical Faculty, University of Zurich, Zurich, Switzerland. Electronic address:

Reliable transportation of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) patient samples from a swabbing station to a diagnostics facility is essential for accurate results. Therefore, cooling or freezing the samples is recommended in case of longer transportation times. In this study, SARS-CoV-2 detectability by RT-PCR was assessed after prolonged unfrozen storage or repetitive freeze-thawing of SARS-CoV-2 samples. SARS-CoV-2-positive patient swabs stored in viral transport medium were exposed to different temperatures (4°C, 25°C, and 35°C) and to repetitive freeze-thawing, to assess the effect of storage conditions on RT-PCR detection. SARS-CoV-2 RNA was still reliably detected by RT-PCR after 21 days of storage in viral transport medium, even when the samples had been stored at 35°C. The maximum observed change in cycle threshold value per day was 0.046 (±0.019) at 35°C, and the maximum observed change in cycle threshold value per freeze-thaw cycle per day was 0.197 (±0.06). Compared with storage at 4°C, viral RNA levels deviated little but significantly when stored at 25°C or 35°C, or after repeated freeze-thawing. The results of this study indicate that viral RNA levels are relatively stable at higher temperatures and repetitive freeze-thawing.
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http://dx.doi.org/10.1016/j.jmoldx.2021.03.003DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7997259PMC
June 2021

Identification of bacteria-derived HLA-bound peptides in melanoma.

Nature 2021 04 17;592(7852):138-143. Epub 2021 Mar 17.

Department of Melanoma Medical Oncology, The University of Texas MD Anderson Cancer Center, Houston, TX, USA.

A variety of species of bacteria are known to colonize human tumours, proliferate within them and modulate immune function, which ultimately affects the survival of patients with cancer and their responses to treatment. However, it is not known whether antigens derived from intracellular bacteria are presented by the human leukocyte antigen class I and II (HLA-I and HLA-II, respectively) molecules of tumour cells, or whether such antigens elicit a tumour-infiltrating T cell immune response. Here we used 16S rRNA gene sequencing and HLA peptidomics to identify a peptide repertoire derived from intracellular bacteria that was presented on HLA-I and HLA-II molecules in melanoma tumours. Our analysis of 17 melanoma metastases (derived from 9 patients) revealed 248 and 35 unique HLA-I and HLA-II peptides, respectively, that were derived from 41 species of bacteria. We identified recurrent bacterial peptides in tumours from different patients, as well as in different tumours from the same patient. Our study reveals that peptides derived from intracellular bacteria can be presented by tumour cells and elicit immune reactivity, and thus provides insight into a mechanism by which bacteria influence activation of the immune system and responses to therapy.
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http://dx.doi.org/10.1038/s41586-021-03368-8DOI Listing
April 2021

Evolution of late-stage metastatic melanoma is dominated by aneuploidy and whole genome doubling.

Nat Commun 2021 03 4;12(1):1434. Epub 2021 Mar 4.

Peter MacCallum Cancer Centre, Melbourne, VIC, Australia.

Although melanoma is initiated by acquisition of point mutations and limited focal copy number alterations in melanocytes-of-origin, the nature of genetic changes that characterise lethal metastatic disease is poorly understood. Here, we analyze the evolution of human melanoma progressing from early to late disease in 13 patients by sampling their tumours at multiple sites and times. Whole exome and genome sequencing data from 88 tumour samples reveals only limited gain of point mutations generally, with net mutational loss in some metastases. In contrast, melanoma evolution is dominated by whole genome doubling and large-scale aneuploidy, in which widespread loss of heterozygosity sculpts the burden of point mutations, neoantigens and structural variants even in treatment-naïve and primary cutaneous melanomas in some patients. These results imply that dysregulation of genomic integrity is a key driver of selective clonal advantage during melanoma progression.
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http://dx.doi.org/10.1038/s41467-021-21576-8DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7933255PMC
March 2021

Oncolytic virotherapy-mediated anti-tumor response: a single-cell perspective.

Cancer Cell 2021 03 21;39(3):394-406.e4. Epub 2021 Jan 21.

Dermatology Department, University Hospital Zurich and Medical Faculty, University of Zurich, 8091 Zurich, Switzerland. Electronic address:

Talimogene laherparepvec (T-VEC) is a genetically modified herpes simplex 1 virus (HSV-1) approved for cancer therapy. We investigate its effect on the clinical, histological, single-cell transcriptomic, and immune repertoire level using repeated fine-needle aspirates (FNAs) of injected and noninjected lesions in primary cutaneous B cell lymphoma (pCBCL). Thirteen patients received intralesional T-VEC, 11 of which demonstrate tumor response in the injected lesions. Using single-cell sequencing of the FNAs, we identify the malignant population and separate three pCBCL subtypes. Twenty-four hours after the injection, we detect HSV-1 transcripts in malignant and nonmalignant cells of the injected lesion but not of the noninjected lesion. Oncolytic virotherapy results in a rapid eradication of malignant cells. It also leads to interferon pathway activation and early influx of natural killer cells, monocytes, and dendritic cells. These events are followed by enrichment in cytotoxic T cells and a decrease of regulatory T cells in injected and noninjected lesions.
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http://dx.doi.org/10.1016/j.ccell.2020.12.022DOI Listing
March 2021

The Tumor Profiler Study: integrated, multi-omic, functional tumor profiling for clinical decision support.

Cancer Cell 2021 03 21;39(3):288-293. Epub 2021 Jan 21.

Roche Pharmaceutical Research and Early Development, Roche Innovation Center Zurich, Wagistrasse 10, 8952 Schlieren, Switzerland.

The application and integration of molecular profiling technologies create novel opportunities for personalized medicine. Here, we introduce the Tumor Profiler Study, an observational trial combining a prospective diagnostic approach to assess the relevance of in-depth tumor profiling to support clinical decision-making with an exploratory approach to improve the biological understanding of the disease.
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http://dx.doi.org/10.1016/j.ccell.2021.01.004DOI Listing
March 2021

Evaluation of the Interplay between the ADAR Editome and Immunotherapy in Melanoma.

Noncoding RNA 2021 Jan 12;7(1). Epub 2021 Jan 12.

Department of Dermatology, University Hospital of Zürich, Gloriastrasse 31, 8091 Zürich, Switzerland.

Background: RNA editing is a highly conserved posttranscriptional mechanism that contributes to transcriptome diversity. In mammals, it includes nucleobase deaminations that convert cytidine (C) into uridine (U) and adenosine (A) into inosine (I). Evidence from cancer studies indicates that RNA-editing enzymes promote certain mechanisms of tumorigenesis. On the other hand, recoding editing in mRNA can generate mutations in proteins that can participate in the Major Histocompatibility Complex (MHC) ligandome and can therefore be recognized by the adaptive immune system. Anti-cancer treatment based on the administration of immune checkpoint inhibitors enhance these natural anti-cancer immune responses.

Results: Based on RNA-Seq datasets, we evaluated the editome of melanoma cell lines generated from patients pre- and post-immunotherapy with immune checkpoint inhibitors. Our results reveal a differential editing in Arthrobacter luteus (Alu) sequences between samples pre-therapy and relapses during therapy with immune checkpoint inhibitors.

Conclusion: These data pave the way towards the development of new diagnostics and therapies targeted to editing that could help in preventing relapses during immunotherapies.
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http://dx.doi.org/10.3390/ncrna7010005DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7838980PMC
January 2021

MITF reprograms the extracellular matrix and focal adhesion in melanoma.

Elife 2021 01 13;10. Epub 2021 Jan 13.

Department of Biochemistry and Molecular Biology, BioMedical Center, Faculty of Medicine, University of Iceland, Reykjavik, Iceland.

The microphthalmia-associated transcription factor (MITF) is a critical regulator of melanocyte development and differentiation. It also plays an important role in melanoma where it has been described as a molecular rheostat that, depending on activity levels, allows reversible switching between different cellular states. Here, we show that MITF directly represses the expression of genes associated with the extracellular matrix (ECM) and focal adhesion pathways in human melanoma cells as well as of regulators of epithelial-to-mesenchymal transition (EMT) such as CDH2, thus affecting cell morphology and cell-matrix interactions. Importantly, we show that these effects of MITF are reversible, as expected from the rheostat model. The number of focal adhesion points increased upon MITF knockdown, a feature observed in drug-resistant melanomas. Cells lacking MITF are similar to the cells of minimal residual disease observed in both human and zebrafish melanomas. Our results suggest that MITF plays a critical role as a repressor of gene expression and is actively involved in shaping the microenvironment of melanoma cells in a cell-autonomous manner.
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http://dx.doi.org/10.7554/eLife.63093DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7857731PMC
January 2021

Blood-based kinase activity profiling: a potential predictor of response to immune checkpoint inhibition in metastatic cancer.

J Immunother Cancer 2020 12;8(2)

Department of Medical Oncology, Oncode Institute, Leiden University Medical Center, Leiden, The Netherlands

Background: Many cancer patients do not obtain clinical benefit from immune checkpoint inhibition. Checkpoint blockade targets T cells, suggesting that tyrosine kinase activity profiling of baseline peripheral blood mononuclear cells may predict clinical outcome.

Methods: Here a total of 160 patients with advanced melanoma or non-small-cell lung cancer (NSCLC), treated with anti-cytotoxic T-lymphocyte-associated protein 4 (anti-CTLA-4) or anti-programmed cell death 1 (anti-PD-1), were divided into five discovery and cross-validation cohorts. The kinase activity profile was generated by analyzing phosphorylation of peripheral blood mononuclear cell lysates in a microarray comprising of 144 peptides derived from sites that are substrates for protein tyrosine kinases. Binary grouping into patients with or without clinical benefit was based on Response Evaluation Criteria in Solid Tumors V.1.1. Predictive models were trained using partial least square discriminant analysis (PLS-DA), performance of the models was evaluated by estimating the correct classification rate (CCR) using cross-validation.

Results: The kinase phosphorylation signatures segregated responders from non-responders by differences in canonical pathways governing T-cell migration, infiltration and co-stimulation. PLS-DA resulted in a CCR of 100% and 93% in the anti-CTLA-4 and anti-PD1 melanoma discovery cohorts, respectively. Cross-validation cohorts to estimate the accuracy of the predictive models showed CCRs of 83% for anti-CTLA-4 and 78% or 68% for anti-PD-1 in melanoma or NSCLC, respectively.

Conclusion: Blood-based kinase activity profiling for response prediction to immune checkpoint inhibitors in melanoma and NSCLC revealed increased kinase activity in pathways associated with T-cell function and led to a classification model with a highly accurate classification rate in cross-validation groups. The predictive value of kinase activity profiling is prospectively verified in an ongoing trial.
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http://dx.doi.org/10.1136/jitc-2020-001607DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7757459PMC
December 2020

Serum CD73 is a prognostic factor in patients with metastatic melanoma and is associated with response to anti-PD-1 therapy.

J Immunother Cancer 2020 12;8(2)

Istituto Nazionale Tumori IRCCS Fondazione Pascale, Napoli, Campania, Italy

Background: Inhibitors of immune checkpoint programmed cell death protein 1 (PD-1) receptor on T cells have shown remarkable clinical outcomes in metastatic melanoma. However, most patients are resistant to therapy. Production of extracellular adenosine, via CD73-mediated catabolism of AMP, contributes to suppress T-cell-mediated responses against cancer. In this study, we analyzed the expression and activity of soluble CD73 in sera of patients with melanoma undergoing anti-PD-1± cytotoxic T-lymphocyte-associated antigen 4 therapy.

Methods: Soluble CD73 expression and activity were retrospectively analyzed in serum of a total of 546 patients with melanoma from different centers before starting treatment (baseline) with anti-PD-1 agents, nivolumab or pembrolizumab, and compared with those of 96 healthy subjects. The CD73 activity was correlated with therapy response and survival of patients.

Results: Patients with melanoma show significantly higher CD73 activity and expression than those observed in healthy donors (p<0.0001). Elevated pretreatment levels of CD73 activity were associated with non-response to therapy with nivolumab or pembrolizumab. During treatment, levels of soluble CD73 activity remain unchanged from baseline and still stratify clinical responders from non-responders. High levels of serum CD73 enzymatic activity associate with reduced overall survival (OS; HR=1.36, 95% CI 1.03 to 1.78; p=0.03) as well as progression-free survival (PFS; HR=1.42, 95% CI 1.13 to 1.79, p=0.003). Further, the multivariate Cox regression analysis indicates that serum CD73 activity is an independent prognostic factor besides serum lactate dehydrogenase levels and the presence of brain metastases for both OS (p=0.009) and PFS (p=0.001).

Conclusion: Our data indicate the relevance of serum CD73 in patients with advanced melanoma receiving anti-PD-1 therapy and support further investigation on targeting CD73 in combination with anti-PD-1 antibodies.
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http://dx.doi.org/10.1136/jitc-2020-001689DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7759961PMC
December 2020

Sustained androgen receptor signaling is a determinant of melanoma cell growth potential and tumorigenesis.

J Exp Med 2021 02;218(2)

Department of Biochemistry, University of Lausanne, Epalinges, Switzerland.

Melanoma susceptibility differs significantly in male versus female populations. Low levels of androgen receptor (AR) in melanocytes of the two sexes are accompanied by heterogeneous expression at various stages of the disease. Irrespective of expression levels, genetic and pharmacological suppression of AR activity in melanoma cells blunts proliferation and induces senescence, while increased AR expression or activation exert opposite effects. AR down-modulation elicits a shared gene expression signature associated with better patient survival, related to interferon and cytokine signaling and DNA damage/repair. AR loss leads to dsDNA breakage, cytoplasmic leakage, and STING activation, with AR anchoring the DNA repair proteins Ku70/Ku80 to RNA Pol II and preventing RNA Pol II-associated DNA damage. AR down-modulation or pharmacological inhibition suppresses melanomagenesis, with increased intratumoral infiltration of macrophages and, in an immune-competent mouse model, cytotoxic T cells. AR provides an attractive target for improved management of melanoma independent of patient sex.
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http://dx.doi.org/10.1084/jem.20201137DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7596884PMC
February 2021

Midkine rewires the melanoma microenvironment toward a tolerogenic and immune-resistant state.

Nat Med 2020 12 19;26(12):1865-1877. Epub 2020 Oct 19.

Melanoma Laboratory, Molecular Oncology Programme, Spanish National Cancer Research Centre (CNIO), Madrid, Spain.

An open question in aggressive cancers such as melanoma is how malignant cells can shift the immune system to pro-tumorigenic functions. Here we identify midkine (MDK) as a melanoma-secreted driver of an inflamed, but immune evasive, microenvironment that defines poor patient prognosis and resistance to immune checkpoint blockade. Mechanistically, MDK was found to control the transcriptome of melanoma cells, allowing for coordinated activation of nuclear factor-κB and downregulation of interferon-associated pathways. The resulting MDK-modulated secretome educated macrophages towards tolerant phenotypes that promoted CD8 T cell dysfunction. In contrast, genetic targeting of MDK sensitized melanoma cells to anti-PD-1/anti-PD-L1 treatment. Emphasizing the translational relevance of these findings, the expression profile of MDK-depleted tumors was enriched in key indicators of a good response to immune checkpoint blockers in independent patient cohorts. Together, these data reveal that MDK acts as an internal modulator of autocrine and paracrine signals that maintain immune suppression in aggressive melanomas.
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http://dx.doi.org/10.1038/s41591-020-1073-3DOI Listing
December 2020

Whole-genome sequencing of acral melanoma reveals genomic complexity and diversity.

Nat Commun 2020 10 16;11(1):5259. Epub 2020 Oct 16.

Center for Rare Melanomas, University of Colorado Cancer Center, Aurora, Colorado, USA.

To increase understanding of the genomic landscape of acral melanoma, a rare form of melanoma occurring on palms, soles or nail beds, whole genome sequencing of 87 tumors with matching transcriptome sequencing for 63 tumors was performed. Here we report that mutational signature analysis reveals a subset of tumors, mostly subungual, with an ultraviolet radiation signature. Significantly mutated genes are BRAF, NRAS, NF1, NOTCH2, PTEN and TYRP1. Mutations and amplification of KIT are also common. Structural rearrangement and copy number signatures show that whole genome duplication, aneuploidy and complex rearrangements are common. Complex rearrangements occur recurrently and are associated with amplification of TERT, CDK4, MDM2, CCND1, PAK1 and GAB2, indicating potential therapeutic options.
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http://dx.doi.org/10.1038/s41467-020-18988-3DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7567804PMC
October 2020

The ALPK1/TIFA/NF-κB axis links a bacterial carcinogen to R-loop-induced replication stress.

Nat Commun 2020 10 9;11(1):5117. Epub 2020 Oct 9.

Institute of Molecular Cancer Research, University of Zurich, 8057, Zurich, Switzerland.

Exposure of gastric epithelial cells to the bacterial carcinogen Helicobacter pylori causes DNA double strand breaks. Here, we show that H. pylori-induced DNA damage occurs co-transcriptionally in S-phase cells that activate NF-κB signaling upon innate immune recognition of the lipopolysaccharide biosynthetic intermediate β-ADP-heptose by the ALPK1/TIFA signaling pathway. DNA damage depends on the bi-functional RfaE enzyme and the Cag pathogenicity island of H. pylori, is accompanied by replication fork stalling and can be observed also in primary cells derived from gastric organoids. Importantly, H. pylori-induced replication stress and DNA damage depend on the presence of co-transcriptional RNA/DNA hybrids (R-loops) that form in infected cells during S-phase as a consequence of β-ADP-heptose/ ALPK1/TIFA/NF-κB signaling. H. pylori resides in close proximity to S-phase cells in the gastric mucosa of gastritis patients. Taken together, our results link bacterial infection and NF-κB-driven innate immune responses to R-loop-dependent replication stress and DNA damage.
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http://dx.doi.org/10.1038/s41467-020-18857-zDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7547021PMC
October 2020

Protein tyrosine phosphatase nonreceptor type 2 controls colorectal cancer development.

J Clin Invest 2021 01;131(1)

Department of Gastroenterology and Hepatology, University Hospital Zurich, and.

Protein tyrosine phosphatase nonreceptor type 2 (PTPN2) recently emerged as a promising cancer immunotherapy target. We set out to investigate the functional role of PTPN2 in the pathogenesis of human colorectal carcinoma (CRC), as its role in immune-silent solid tumors is poorly understood. We demonstrate that in human CRC, increased PTPN2 expression and activity correlated with disease progression and decreased immune responses in tumor tissues. In particular, stage II and III tumors displayed enhanced PTPN2 protein expression in tumor-infiltrating T cells, and increased PTPN2 levels negatively correlated with expression of PD-1, CTLA4, STAT1, and granzyme A. In vivo, T cell- and DC-specific PTPN2 deletion reduced tumor burden in several CRC models by promoting CD44+ effector/memory T cells, as well as CD8+ T cell infiltration and cytotoxicity in the tumor. In direct relevance to CRC treatment, T cell-specific PTPN2 deletion potentiated anti-PD-1 efficacy and induced antitumor memory formation upon tumor rechallenge in vivo. Our data suggest a role for PTPN2 in suppressing antitumor immunity and promoting tumor development in patients with CRC. Our in vivo results identify PTPN2 as a key player in controlling the immunogenicity of CRC, with the strong potential to be exploited for cancer immunotherapy.
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http://dx.doi.org/10.1172/JCI140281DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7773375PMC
January 2021

Retrospective Analysis of Treatment and Complications of Immune Checkpoint Inhibitor-Associated Colitis: Histological Ulcerations as Potential Predictor for a Steroid-Refractory Disease Course.

Inflamm Intest Dis 2020 Aug 26;5(3):109-116. Epub 2020 May 26.

Division of Gastroenterology and Hepatology, University Hospital Zurich, University of Zurich, Zurich, Switzerland.

Background/aims: Among the severe immune-related adverse events (irAEs) that occur with immune checkpoint inhibitor (ICI) therapy, colitis is the most frequent one. This study aimed at describing the experience from the largest gastroenterology unit in Switzerland with immune checkpoint inhibitor-associated colitis (ICIAC), its clinical presentation, management, and outcomes.

Methods: We performed a retrospective review of patients who were referred for the evaluation of ICIAC between January 2011 and October 2018 to the Division of Gastroenterology and Hepatology, University Hospital Zurich.

Results: Thirty-three patients with immune-related colitis grade 3 or 4 met the inclusion criteria and were analyzed in detail: All patients had diarrhea, 64% had abdominal pain, 42% had bloody stool, 27% had emesis, and 18% developed fever. In total, 33% were successfully treated with corticosteroids alone; 66% were steroid-refractory and treated with infliximab or vedolizumab. Two of these patients developed severe complications requiring surgery. All patients reached complete remission of ICIAC and its symptoms. At colonoscopy, ulcerations were seen in 37% of steroid-refractory versus 63% of steroid-responsive cases. Deep histological ulcerations invading the submucosa were only present in steroid-refractory cases.

Conclusion: ICIAC is a severe irAE which frequently requires high-dose steroids and a close follow-up due to deleterious complications. The detection of histologically diagnosed deep ulcerations may predict a steroid-refractory course and may warrant early application of infliximab. However, larger studies are required to confirm our findings.
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http://dx.doi.org/10.1159/000507579DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7506225PMC
August 2020

NRAS melanoma tumor formation is reduced by p38-MAPK14 activation in zebrafish models and NRAS-mutated human melanoma cells.

Pigment Cell Melanoma Res 2021 03 24;34(2):150-162. Epub 2020 Sep 24.

University Hospital Zurich, University of Zurich, Zurich, Switzerland.

Oncogenic BRAF and NRAS mutations drive human melanoma initiation. We used transgenic zebrafish to model NRAS-mutant melanoma, and the rapid tumor onset allowed us to study candidate tumor suppressors. We identified P38α-MAPK14 as a potential tumor suppressor in The Cancer Genome Atlas melanoma cohort of NRAS-mutant melanomas, and overexpression significantly increased the time to tumor onset in transgenic zebrafish with NRAS-driven melanoma. Pharmacological activation of P38α-MAPK14 using anisomycin reduced in vitro viability of melanoma cultures, which we confirmed by stable overexpression of p38α. We observed that the viability of MEK inhibitor resistant melanoma cells could be reduced by combined treatment of anisomycin and MEK inhibition. Our study demonstrates that activating the p38α-MAPK14 pathway in the presence of oncogenic NRAS abrogates melanoma in vitro and in vivo. SIGNIFICANCE: The significance of our study is in the accountability of NRAS mutations in melanoma. We demonstrate here that activation of p38α-MAPK14 pathway can abrogate NRAS-mutant melanoma which is contrary to the previously published role of p38α-MAPK14 pathway in BRAF mutant melanoma. These results implicate that BRAF and NRAS-mutant melanoma may not be identical biologically. We also demonstrate the translational benefit of our study by using a small molecule compound-anisomycin (already in use for other diseases in clinical trials) to activate p38α-MAPK14 pathway.
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http://dx.doi.org/10.1111/pcmr.12925DOI Listing
March 2021

Multi-site clonality analysis uncovers pervasive heterogeneity across melanoma metastases.

Nat Commun 2020 08 27;11(1):4306. Epub 2020 Aug 27.

Experimental Cancer Genetics, The Wellcome Sanger Institute, Hinxton, Cambridgeshire, UK.

Metastatic melanoma carries a poor prognosis despite modern systemic therapies. Understanding the evolution of the disease could help inform patient management. Through whole-genome sequencing of 13 melanoma metastases sampled at autopsy from a treatment naïve patient and by leveraging the analytical power of multi-sample analyses, we reveal evidence of diversification among metastatic lineages. UV-induced mutations dominate the trunk, whereas APOBEC-associated mutations are found in the branches of the evolutionary tree. Multi-sample analyses from a further seven patients confirmed that lineage diversification was pervasive, representing an important mode of melanoma dissemination. Our analyses demonstrate that joint analysis of cancer cell fraction estimates across multiple metastases can uncover previously unrecognised levels of tumour heterogeneity and highlight the limitations of inferring heterogeneity from a single biopsy.
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http://dx.doi.org/10.1038/s41467-020-18060-0DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7453196PMC
August 2020

Toxicity of combined targeted therapy and concurrent radiotherapy in metastatic melanoma patients: a single-center retrospective analysis.

Melanoma Res 2020 12;30(6):552-561

Department of Dermatology, University Hospital of Zurich, University of Zurich, Gloriastrasse.

The Eastern Cooperative Oncology Group consensus guidelines from 2016 recommend interruption of targeted therapy with BRAF- and MEK-inhibitors during radiotherapy with data being based mostly on BRAF monotherapy. The aim of this study is to provide data on the safety of concurrent radiotherapy and combination targeted therapy with BRAF- and MEK-inhibitors. A total of 32 patients with 51 sessions of radiotherapy from one center receiving concurrent radiotherapy and BRAF- and MEK- inhibitors were included. Radiotherapy-associated toxicities were retrospectively collected. Incidence was compared between three groups: (A) targeted therapy during radiotherapy with and, (B) without interruption, and (C) radiotherapy before the start of targeted therapy. Survival and local disease control were examined. Targeted therapy was interrupted during radiotherapy in 16, not interrupted in 14, and only started after radiotherapy in 21 sessions. Stereotactic radiotherapy was applied in 28 sessions, conventionally fractionated radiotherapy in 23. The brain was the most common site of irradiation (n = 36). Radiotherapy-associated toxicities occurred in 41.2% (n = 21) of sessions and did not differ significantly among the groups. Overall survival was 11.7 months and progression-free survival was 8.4 months. No increase in radiotherapy-associated toxicity was seen where combination targeted therapy was not interrupted during radiotherapy. Prospective clinical trials are warranted to support our findings.
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http://dx.doi.org/10.1097/CMR.0000000000000682DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7643789PMC
December 2020
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